Ancient Recipe May Burst the MRSA Bubble

It reads like an act from Shakespeare or a scene from Harry Potter. A 1000-year-old book is opened, an ancient tongue is translated, and a magical potion is brewed. The potion is tested against modern-day organisms, and it works!

Researchers from the University of Nottingham in the United Kingdom recently translated a 10th-century, Old English recipe for an eye salve from Bald’s Leechbook. Its ingredients included 2 species of Allium (garlic, onion, and/or leek), wine, and oxgall (cow stomach bile). The mixture would then sit in a brass cauldron for 9 days, after which it could be strained and used as a topical salve for eye ailments.

Since many of the ailments described in Bald’s Leechbook are bacterial in origin, the researchers hypothesized that the treatments may possess some antimicrobial activity. So, they brewed 4 batches of the salve, each time from scratch, as well as a “placebo” consisting of distilled water and a brass sheet, minus the food components. Using collagen coated with methicillin-resistant Staphylococcus aureus (MRSA) to mimic wound infections, they tested each individual ingredient and the salve batches to determine whether any of them inhibited bacterial growth.

While none of the individual ingredients showed efficacy, the full recipe drastically decreased colony counts. Perhaps even more exciting, the salve was found to be active against biofilm produced by the bacteria, which is something only a handful of current antimicrobials can do.

Additional in vivo testing was carried out in MRSA wounds in animal models at Texas Tech University. The mouse models confirmed the initial finding, as up to 90% of MRSA in their wounds was killed. The researchers commented that the salve performed “as good if not better than the conventional antibiotics we used.”^1-2

Interestingly, none of the individual components demonstrated any antimicrobial activity. All of the ingredients have been studied and reported to inhibit bacterial growth in the past; however, the exact concentration or amount is unknown. What, if any, chemical reaction takes place during the 9-day curing period is also yet to be determined.

The Allium family of plants includes garlic, Chinese leek, and onion. The antimicrobial benefits of Allium plants have been attributed to the presence of sulphides, most notably diallyl sulphide (DAS) and diallyl disulphide (DADS). Several in vitro studies have compared the antimicrobial activity of Allium oils and extracts with commonly used antibiotics.

One study tested Allium extracts against methicillin, penicillin, cefotaxime, and tetracycline in S. aureus and MRSA isolates. Overall, researchers found that garlic extracts contain higher sulphide percentages than Chinese leek extracts, therefore showing more activity. The antibiotics demonstrated lower minimum inhibitory concentration (MIC) values against S. aureus than the Allium extracts (0.5 mcg/mL to 1 mcg/mL for antibiotics versus 0.5 mcg/mL to 36 mcg/mL for Allium). However, the Allium extracts showed lower MIC values against the resistant MRSA strain (>64 mcg/mL for antibiotics versus 2 mcg/mL to 48 mcg/mL for Allium). The researchers commented that the more disulphide bonds present, the more potent the anti-staphylococcal activity.³

The same investigators then tested their theories in mice models of S. aureus infection. Mice were infected with MRSA and orally administered placebo, garlic oil, DAS, DADS, or vancomycin 16 hours after infection. At 24 hours post-infection (8 hours post-treatment administration), the mice were sacrificed and plasma, liver, spleen, and kidney samples were obtained. Compared with placebo and vancomycin, all 3 Allium compounds significantly reduced MRSA colonies in plasma, liver, kidney, and spleen. As previously found, the higher the concentration of sulphide administered, the greater the decrease in bacteria load. The authors also said there appears to be an upper limit of toxicity for these compounds, so additional animal studies must be conducted to find the optimal concentration to balance efficacy and toxicity.⁴

Alcohol, or more specifically, ethanol, has long been used as an antimicrobial agent. In concentrations >60%, ethanol is routinely used in health care settings as a hand sanitizer in lieu of washing with soap and water.⁵ Additionally, ethanol locks (10% to 70%) have been used to treat indwelling catheter-related infections when removal of the catheter is not possible.⁶ Ethanol has activity against both Gram-positive and Gram-negative bacteria.

Oxgall is a bit harder to explain. While studies on oxgall and staphylococci are lacking, there are some reports of oxgall’s effect on other Gram-positive bacteria. One in vitro study reported that oxgall reduced the adhesion ability of Enterococci.⁷ Another study reported that the addition of oxgall increased susceptibilities to cell wall active antibiotics in bifidobacteria (e.g., Lactobacillus).⁸

Whether the same effects are exhibited in staphylococci are left to be seen. Then again, perhaps Bald was just looking for an alternative to “eye of newt”?

The brass vessel, on the other hand, is easier to explain. Brass is a metal made up of mostly zinc and copper, the latter of which has long been recognized as possessing antimicrobial activity.

A recent mouse model explored the ability of copper to aid in wound healing. Mice were randomized to receive copper-impregnated wound dressing, silver-impregnated wound dressing, or an identical wound dressing without copper or silver. The mice used in the study were a specific type of diabetic mouse, chosen for their inability to heal wounds properly. The copper-treated mice not only healed faster than the silver- or placebo-treated mice in regards to wound size, but also demonstrated time to healing similar to that of non-diabetic, non-healing-impaired mice.⁹

Another study found that copper also had the ability to penetrate biofilm. Researchers tested vancomycin, chloramphenicol, and several copper compounds against biofilm produced by a strain of MRSA. Vancomycin concentrations at 400 x MIC were required to get some biofilm reduction (44%). However, copper concentrations at only 3 x MIC were required to reduce the biofilm by 68%. Although the researchers concluded that copper was more effective than vancomycin in reducing biofilm, they commented that copper can also be cytotoxic, though the optimal copper compound and concentration has yet to be determined.¹⁰

This is a fascinating story of discovering a viable and potent remedy from 1000 years ago. The text is translated from Old English, which makes today’s prescribers’ handwriting seem legible. As the name implies, the first MRSA isolate was not identified until 1960, after the development of methicillin. Finding an ancient compound that is active against a modern-day resistance mechanism is astounding, and the amount of attention that this discovery has generated for the field of antibiotic discovery is priceless.

The University of Nottingham researchers are slated to present their findings at an annual microbiology conference in the United Kingdom. With further investigation, perhaps some of these questions may be answered.

References:

1. Rayner E. AncientBiotics — a medieval remedy for modern day superbugs? The University of Nottingham. Available at: http://www.nottingham.ac.uk/news/pressreleases/2015/march/ancientbiotics---a-medieval-remedy-for-modern-day-superbugs.aspx [Accessed 4/2/2015]

2. Harrison F, Roberts A, Rumbaugh K, Lee C, Diggle SP. A 1,000-year-old antimicrobial remedy with anti-staphylococcal activity. [absctract] Society for General microbiology Annual Conference 2015. Birmingham, UK.

3. Tsao SM, Yin MC. In-vitro antimicrobial activity of four diallyl sulphides occurring naturally in garlic and Chinese leek oils. J Med Microbiol. 2001;50:646-9.

4. Tsao SM, Hsu CC, Yin MC. Garlic extract and two diallyl sulphides inhibit methicillin-resistant Staphylococcus aureus infection in BALB/cA mice. J Antimicrob Chemother. 2003;52:974-80.

5. Guideline for hand hygiene in health-care settings. MMWR. 2002;51:RR-16.

6. Mermel LA, Allon M, Bouza E, et al. Clinical practice guidelines for the diagnosis and management of intravascular catheter-related infection: 2009 update by the Infectious Diseases Society of America. Clin Infect Dis. 2009;49:1-45.

7. Marcinakova M, Klingberg TD, Laukova A, Budde BB. The effect of pH, bile and calcium on the adhesion ability of probiotic enterococci of animal origin to the porcine jejunal epithelial cell line IPEC-J2. Anaerobe. 2010;2:120-4.

8. Kheadr E, Dabour N, Le Lay C, Lacroix C, Fliss I. Antibiotic susceptibility profile of bifidobacteria as affected by oxgall, acid, and hydrogen peroxide stress. Antimicrob Agents Chemother. 2007;51:169-74.

9. Borkow G, Gabbay J, Dardik R, et al. Molecular mechanisms of enhanced wound healing by copper oxide-impregnated dressings. Wound Rep Reg. 2010;18:266-75.

10. Beeton ML, Aldrich-Wright JR, Bolhuis A. The antimicrobial and antibiofilm activities of copper(II) complexes. J Inorg Biochem. 2014;140:167-72.